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ReviewCite this article: Lane N. 2015 The unseenworld: reflections on Leeuwenhoek (1677)

‘Concerning little animals’. Phil. Trans. R. Soc. B

370: 20140344.http://dx.doi.org/10.1098/rstb.2014.0344

One contribution of 18 to a theme issue

‘Celebrating 350 years of Philosophical

Transactions: life sciences papers’.

Subject Areas:microbiology

Keywords:Leeuwenhoek, animalcule, protozoa,

bacteria, eukaryote, tree of life

Author for correspondence:Nick Lane

e-mail: [email protected]

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The unseen world: reflections onLeeuwenhoek (1677) ‘Concerninglittle animals’

Nick Lane

Department of Genetics, Evolution and Environment, University College London, London WC1E 6BT, UK

Leeuwenhoek’s 1677 paper, the famous ‘letter on the protozoa’, gives the firstdetailed description of protists and bacteria living in a range of environments.The colloquial, diaristic style conceals the workings of a startlingly originalexperimental mind. Later scientists could not match the resolution and clarityof Leeuwenhoek’s microscopes, so his discoveries were doubted or even dis-missed over the following centuries, limiting their direct influence on thehistory of biology; but work in the twentieth century confirmed Leeuwenhoek’sdiscovery of bacterial cells, with a resolution of less than 1 mm. Leeuwenhoekdelighted most in the forms, interactions and behaviour of his little ‘animal-cules’, which inhabited a previously unimagined microcosmos. In thesereflections on the scientific reach of Leeuwenhoek’s ideas and observations,I equate his questions with the preoccupations of our genomic era: whatis the nature of Leeuwenhoek’s animalcules, where do they come from,how do they relate to each other? Even with the powerful tools of modernbiology, the answers are far from resolved—these questions still challengeour understanding of microbial evolution. This commentary was written tocelebrate the 350th anniversary of the journal Philosophical Transactions of theRoyal Society.

My work, which I’ve done for a long time, was not pursued in order to gain the praiseI now enjoy, but chiefly from a craving after knowledge, which I notice resides in memore than most other men.

Leeuwenhoek, Letter of 12 June 1716

Leeuwenhoek is universally acknowledged as the father of microbiology. Hediscovered both protists and bacteria [1]. More than being the first to see thisunimagined world of ‘animalcules’, he was the first even to think of look-ing—certainly, the first with the power to see. Using his own deceptivelysimple, single-lensed microscopes, he did not merely observe, but conductedingenious experiments, exploring and manipulating his microscopic universewith a curiosity that belied his lack of a map or bearings. Leeuwenhoek(figure 1) was a pioneer, a scientist of the highest calibre, yet his reputation suf-fered at the hands of those who envied his fame or scorned his unschooledorigins, as well as through his own mistrustful secrecy of his methods, whichopened a world that others could not comprehend. The verification of thisnew world by the natural philosophers of the nascent Royal Society laid outthe ground rules that still delineate science today, but the freshness andwonder, the sheer thrill of Leeuwenhoek’s discoveries, transmit directlydown the centuries to biologists today. Microbiologists and phylogeneticistscontinue to argue about the nature of Leeuwenhoek’s little animals, if inmore elaborate terms. Only now are we beginning to find answers—and sur-prisingly uncertain answers—to the questions that drove Leeuwenhoek:where did this multitude of tiny ‘animals’ come from, why such variety insize and behaviour; how to distinguish and classify them?

Leeuwenhoek’s 1677 paper [1] was not his first contribution to PhilosophicalTransactions, nor was it his first mention of little animals living in water.

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Figure 1. Portrait of Leeuwenhoek by Jan Verkolje, 1686, at age 54. Copyright & The Royal Society.

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The paper was translated (‘English’d’) from low Dutch,and excerpted to half its original length by the redoubtableHenry Oldenburg, first Secretary of the Royal Society andfounding editor of Philosophical Transactions. Oldenburgcorresponded so widely across Europe that he was imprisonedin the Tower for suspected espionage in 1667, during theSecond Anglo-Dutch War (when the Dutch colony of NewAmsterdam was renamed New York). Oldenburg lateradopted a pseudonym, the anagram ‘Grubendol’, to avertsuspicion (it would arouse mine). Among his regular Dutchcorrespondents were the surgeon Regnerus de Graaf andstatesman Constantijn Huygens, father of famed astrono-mer Christiaan Huygens, both of whom wrote epistles toOldenburg introducing ‘the exceedingly curious and indus-trious’ Leeuwenhoek, Huygens adding the helpful note‘or Leawenhook, according to your orthographie’ [2]. Thatwas in 1673; by 1677, Leeuwenhoek was well known to the

Royal Society, but by no means were his reports acceptedon trust.

Oldenburg published several of Leeuwenhoek’s letters in1673 and 1674, which dealt with interesting but uncontentiousmatters, such as the structure of the bee sting. Equivalentmicroscopic structures of objects visible to the naked eye hadbeen illuminated by Robert Hooke in his Micrographia nearlya decade earlier; indeed, it is to Hooke that we owe the word‘cell’, which he used to denote the boxy spaces (reminiscentof the small rooms in a monastery) that make up the structureof cork [3]. From some of Leeuwenhoek’s slightly waspishremarks in his early letters, he had almost certainly seena copy of Micrographia on his visit to London in 1667 or1668, when the book was practically a fashion accessory(‘the most ingenious book I read in all my life’, wrote Pepys,who stayed up all night with it; Pepys reputedly stayed upall night often, though rarely with a book). Leeuwenhoek

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Figure 2. First and last pages of Leeuwenhoek’s 1676 letter to Oldenburg, in the hand of a copyist. Copyright & The Royal Society.


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first courted controversy in a letter of September 1674.Describing a nearby lake, Berkelse Mere, he noted that itswater was very clear in winter ‘but at the beginning ormiddle of summer it becomes whitish, and there are thenlittle green clouds floating in it’ [4]. These clouds containedwispy ‘green streaks, spirally wound serpent-wise, and orderlyarranged’—the beautiful green alga Spirogyra. Then came Leeu-wenhoek’s first mention of little animals: ‘among these streaksthere were besides very many little animalcules . . . And themotion of most of these animalcules in the water was soswift, and so various upwards, downwards and round aboutthat ‘twas wonderful to see: and I judged that some of theselittle creatures were above a thousand times smaller than thesmallest ones I have ever yet seen upon the rind of cheese’(by which he meant mites) [4].

Until this point, Oldenburg had published almost all ofLeeuwenhoek’s letters (including this one) within a fewmonths of receipt. Now, he drew pause. Of the next 12 letterssent by Leeuwenhoek, only three were published, and nonethat touched on animalcules. Oldenburg had every reasonto be suspicious; as Leeuwenhoek wrote to Hooke a fewyears later ‘I suffer many contradictions and oft-times hearit said that I do but tell fairy tales about the little animals’[5]. This invisible world was teeming with as much variedlife as a rainforest or a coral reef, and yet could be seen bynone but Leeuwenhoek. No wonder Oldenburg and hiscolleagues had doubts. Set against this background, Leeu-wenhoek wrote his eighteenth letter to the Royal Society,dated October 1676, the celebrated ‘letter on the protozoa’,

which Oldenburg excerpted, translated and published asthe 1677 paper. It opens with a bang: ‘In 1675 I discoveredliving creatures in Rain water which had stood but fewdays in a new earthen pot, glased blew [i.e. painted blue]within. This invited me to view this water with great atten-tion, especially those little animals appearing to me tenthousand times less than those represented by Mons.Swamerdam and called by him Water fleas or Water-lice,which may be perceived in the water with the naked eye’ [1].

Clifford Dobell, in his delightful biography published in1932 (300 years after Leeuwenhoek’s birth), notes that Olden-burg’s translation is good but not perfect [6]. That’s notsurprising. While Oldenburg knew the language, he had noknowledge of the organisms themselves. In contrast, Dobellwas a distinguished microbiologist, a Fellow of the RoyalSociety, and had the great benefit of hindsight. His biographywas a labour of love, written over 25 years, frequently in themiddle of the night, while carrying out his own research onintestinal protozoa and other protists. Dobell taught himselfDutch and translated Leeuwenhoek’s letters painstakingly—written, as they were, in a colloquial Dutch no longer in use,and in the beautiful but scarcely legible hand of a copyist(figure 2). Dobell revelled in the precise beauty of Leeuwen-hoek’s descriptions of Euglena, Vorticella and many otherprotists and bacteria, which leapt off the page, immediatelyrecognizable to this expert kindred spirit. Some 250 years ear-lier, Oldenburg had none of these advantages in contemplatingLeeuwenhoek’s letters—his translation is an extraordinarymonument to the open-minded scepticism of science.



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I sketch this background because the paper itself is unu-sual even in Leeuwenhoek’s oeuvre, taking the form of adiary. On a cursory reading, it seems almost embarrassinglynaive to the modern ear—the earthen pot ‘glased blewwithin’ in the first sentence is a good example (but see [7]for a discussion of Dutch prose style in the seventeenth cen-tury). We learn that on ‘the 17th of this month of June itrained very hard; and I catched some of that rain water in anew Porcelain dish, which had never been used before, butfound no living creatures at all in it’ [1]. On it goes, with pre-cise but apparently irrelevant details. ‘In the open Court ofmy house I have a well, which is about 15 foot deep, beforeone comes to the water. It is encompassed with high walls,so that the Sun, though in Cancer, yet can hardly shinemuch upon it. This water comes out of the ground, whichis sandy, with such a power, that when I have laboured toempty the well, I could not so do it but there remainedever a foots depth of water in it. This water is in Summertime so cold, that you cannot possibly endure your hand init for any reasonable time’ [1]. And my favourite: ‘July 271676. I went to the sea-side, at Schlevelingen, the windcoming from the Sea with a very warm Sun-shine; and view-ing some of the Sea-water very attentively, I discovereddivers living animals therein. I gave to a man, that wentinto the Sea to wash himself, a new glass-bottle, boughton purpose for that end, intreating him, that being on theSea, he would first wash it well twice, or thrice, and thenfill it full of the Sea-water; which desire of mine havingbeen complied with, I tyed the bottle close with a cleanbladder’ [1].

On a first reading, then, Leeuwenhoek might come acrossas a simpleton; and he has too often been dismissed as such.One can only smile at the image of Leeuwenhoek on thebeach, pressing his pre-prepared bottles onto strangers. Butwhich details are important? How should he have chartedthis abundant new world? We need to appreciate severalpoints. This letter was intended to defend his discoveries—‘merely so as to make my observations more credible inEngland and elsewhere’ [8]. Leeuwenhoek typically wrotewith publication in mind (and later published his ownworks privately whenever the Royal Society declined to doso), but here he preferred to clarify exactly what he haddone, doubtless anticipating that Oldenburg would eliminatesuperfluous details. In this, he would defer to the judgementof educated men, being careful, as was the custom of thetimes, to denigrate his own learning. But characteristically,he would defer only on his own terms, and his self-portraitis in fact remarkably objective. ‘I have oft-times beenbesought, by divers gentlemen, to set down on paper whatI have beheld through my newly invented Microscopia: but Ihave generally declined; first, because I have no style, orpen, wherewith to express my thoughts properly; second,because I have not been brought up to languages or arts,but only to business; and in the third place, because I donot gladly suffer contradiction or censure from others’ [9].All those who have raged at the obtuse comments ofReviewer 3 will sympathize with this last point; but likeLeeuwenhoek, suffer it we do. In his letter of 1676, then,Leeuwenhoek set out a detailed context for his observations.Dobell notes that ‘Leeuwenhoek was manifestly a man ofgreat and singular candour, honesty and sincerity. He wasreligiously plain and straightforward in all he did, and there-fore sometimes almost immodestly frank in describing his

observations. It never occurred to him that Truth couldappear indecent’ [10].

On a closer reading, the colloquial manner of Leeuwen-hoek’s letter conceals the workings of his precise andmethodical mind. Leeuwenhoek was acutely aware ofcontamination; he replenished evaporated water with snow-water, the purest then available, making every effort not tointroduce little animals from any other source. He sampledwater from many different sources—his well, the sea, rainwater, drain pipes, lakes—always taking care to clean hisreceptacles. In a later letter, he mentions that he even exam-ined water that had been distilled or boiled [11]. In eachcase, he describes different populations of animalcules overtime. Time is critical. Frequently, he observes nothing for aweek, checking each day, before reporting a profusion oflittle animals of diverse types, replicating themselves overseveral days before dying back again. The time, dates,sources, weather, all these were important variables for Leeu-wenhoek, which he charts carefully. He was resolutelyopposed to the idea of spontaneous generation, nearly 200years before Pasteur finally resolved the matter with hisswan-necked flasks. Leeuwenhoek later described the pro-creation of cells via copulation or schism to releasedaughter cells in arresting detail. But his early disbelief ofspontaneous generation is implicit in the comparisons ofhis 1677 paper, in his care to avoid contamination, and hisestimation of rates of growth.

Leeuwenhoek also reports experiments, adding pepper-corns to water, both crushed and uncrushed (as wellas ginger, cloves, nutmeg and vinegar, omitted fromOldenburg’s excerpts for Philosophical Transactions). In theseinfusions, Leeuwenhoek observed an astonishing prolifer-ation of tiny animals ‘incredibly small; nay, so small, in mysight, that I judged that even if 100 of these very wee animalslay stretched out one against another, they could not reachthe length of a grain of course sand; and if this be true,then ten hundred thousand of these living creatures couldscarce equal the bulk of a course grain of sand’ [1]. Again,the colloquial language deceives. In a clarification sent toConstantijn Huygens and Hooke, Leeuwenhoek writes‘Let’s assume that such a sand-grain is so big, that 80 ofthem, lying one against the other, would make up thelength of one inch’ [12]. He goes on to calculate the numberof animalcules in a cubic inch; for our purposes here, hiscalculation puts the length of his ‘very wee animals’ at lessthan 3 mm. Bacteria. (He later describes bacterial motilityunequivocally [13]). He also notes that he deliberately under-estimates the number of bacteria in a drop of water—‘for thereason that the number of animalcules in so small a quantityof water would else be so big, that ‘twould not be credited:and when I stated in my letter of 9th October 1676, thatthere were upwards of 1 000 000 living creatures in onedrop of pepper-water, I might with truth have put thenumber at eight times as many’ [14]. An innocent, earlyexample of spinning data to sell to a journal?

But the natural philosophers of the Royal Society, in pio-neering the methods we still use in science today, were noteasily spun. Leeuwenhoek’s letter had been read aloud overseveral sessions and attracted great interest, verging on con-sternation. Oldenburg wrote to Leeuwenhoek, asking himto ‘acquaint us with his method of observing, that othersmay confirm such Observations as these’, and to providedrawings [15]. Leeuwenhoek declined, throughout his


(a) (b)

Figure 3. (a) Rotifers, hydra and vorticellids associated with a duckweed root, from a Delft canal. From Leeuwenhoek [16]. (b) Bacteria from Leeuwenhoek’s mouth;the dotted line portrays movement. From Leeuwenhoek [17]. Copyright & The Royal Society.


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life, to give any description of his microscopical methods (‘forreasons best known to himself’, said Hooke; though sciencehas hardly resolved the issue of intellectual property sincethen). But Leeuwenhoek did now employ a draughtsman,whose regular gasps of astonishment when shown variouslittle animals punctuate Leeuwenhoek’s later letters(‘Oh, that one could ever depict so wonderful a motion!’).Some of these limner’s drawings are shown in figure 3.Leeuwenhoek also sent eight testimonies from gentlemenof repute—a Lutheran minister, a notary and a barrister,among others. It is striking to the modern reader that noneof these gentlemen were natural philosophers acquaintedwith the methods of science; but according to the historianSteven Shapin, it was the bond of the gentleman thatcounted. The practice of signed testimonies from gentlemenwas common in the seventeenth century; the fact that Leeu-wenhoek called upon eight such testimonies attests to theunprecedented character of his findings, but also perhaps tohis lower social standing [18].

No doubt all this was helpful, but it was counteredby letters from others such as Christiaan Huygens (son ofConstanijn), then in Paris, who at that time remained scepti-cal, as was his wont: ‘I should greatly like to know how muchcredence our Mr Leeuwenhoek’s observations obtain amongyou. He resolves everything into little globules; but for mypart, after vainly trying to see some of the things which hesees, I much misdoubt me whether they be not illusions ofhis sight’ [19]. The Royal Society tasked Nehemiah Grew,the botanist, to reproduce Leeuwenhoek’s work, but Grewfailed; so in 1677, on succeeding Grew as Secretary, Hookehimself turned his mind back to microscopy. Hooke tooinitially failed, but on his third attempt to reproduceLeeuwenhoek’s findings with pepper-water (and otherinfusions), Hooke did succeed in seeing the animalcules—‘some of these so exceeding small that millions of millionsmight be contained in one drop of water’ [20] (actually farless precise than Leeuwenhoek). He went on to write ‘Itseems very wonderful that there should be such an infinite

number of animalls in soe imperceptible quantity of matter.That these animalls should be soe perfectly shaped andindeed with such curious organs of motion as to be ableto move nimbly, to turne, stay, accelerate and retard theirprogresse at pleasure. And it was not less surprising tofind that these were gygantick monsters [protozoa] in com-parison of a lesser sort which almost filled the water[bacteria]’ [21].

Unlike Leeuwenhoek, Hooke gave precise details of hismicroscopical methods, and demonstrated them before thegathered fellows, including Sir Christopher Wren, later pub-lishing both his methods and observations in Microscopium(1678) [20]. He even taught himself Dutch, so that he couldread the letters of the ‘ingenious Mr Leeuwenhoek’. Asnoted by the microscopist Brian J. Ford [22] and microbiologistHoward Gest [23], Hooke was a central and too-often over-looked figure in the history of microbiology: his earlier bookMicrographia (1665) most likely inspired Leeuwenhoek tobegin his own microscopical studies. Without Hooke’s sup-port and verification—a task beyond several of the bestmicroscopists of the age, including Grew—Leeuwenhoekmight easily have been dismissed as a charlatan. Instead,through Hooke’s impressive demonstrations, and withthe direct support of the patron of the Royal Society, KingCharles II, Leeuwenhoek was elected a Fellow in 1680.Others had independently changed their view of Leeuwen-hoek in the interim, but that did little to alter the course ofevents. Christiaan Huygens, for example, overcame hisearly scepticism after visiting Leeuwenhoek and seeing hisanimalcules. He went on to grind his own lenses, observingvarious protists himself [24]. Indeed, Huygens made anumber of pioneering observations, but these remained inmanuscript and were unpublished until the turn of thetwentieth century [25].

Ironically, Hooke’s admirable comments on the construc-tion of microscopes might have undermined Leeuwenhoek’slater reputation. Hooke made various types of microscope.He much preferred using larger instruments with two


(a) (b)

(c) (d)

Figure 4. (a) Replica of a single-lens microscope by Leeuwenhoek (Image by Jeroen Rouwkema. Licensed under CC BY-SA 3.0 via Wikimedia Commons). (b,d)Photomicrographs taken using simple single-lens microscopes including one of Leeuwenhoek’s originals in Utrecht, by Brian Ford (Copyright & Brian J. Ford). (b) Anair-dried smear of Ford’s own blood through the original van Leeuwenhoek microscope at Utrecht, showing red blood cells and a granulocyte with its lobed nucleus(upper right; about 2 mm in diameter). (c) Spiral bacteria (Spirillum volutans) imaged through a replica microscope with a lens ground from spinel; each bacterialcell is about 20 mm in length. (d ) The intestinal protist parasite Giardia intestinalis imaged through a replica soda-glass produced by Brian Ford [28,29].


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lenses, but in the Preface to Micrographia [26] he also describedhow to make ‘simple’ microscopes with a single lens—what became known as a Leeuwenhoek microscope [27].The lens is produced by melting Venice glass into thin threads,containing little globules, which are then ground andpolished, and mounted against a needle hole pricked througha thin plate of brass (figure 4). ‘If . . . an Object, plac’dvery near, be look’d at through it, it will both magnifie andmake some objects more distinct than any of the great Micro-scopes. But because these, though exceeding easily made,are yet very troublesome to be us’d, because of their smallness,and the nearness of the Object; therefore to prevent both ofthese, and yet have only two refractions, I provided me aTube of Brass’ [26]. In 1678, Hooke reiterated his dislike ofsingle-lens microscopes: ‘I have found the use of them offen-sive to my eye, and to have much strained the sight, whichwas the reason why I omitted to make use of them, thoughin truth they make the object appear much more clear and dis-tinct, and magnifie as much as the double Microscopes: nay tothose whose eyes can well endure it, ‘tis possible with a singleMicroscope to make discoveries much better than with adouble one, because the colours which do much disturbthe clear vision in double Microscopes is clearly avoided andprevented with the single’ [20].

It seems that Hooke’s aversion to simple single-lensmicroscopes passed on down the generations, but not his

appreciation of their merits. The compound microscope, withits refractive aberrations, became the tool of choice, andLeeuwenhoek’s microscopes were quietly forgotten, theiroblivion hastened by Leeuwenhoek’s own secrecy, notwith-standing his gift of 13 microscopes, with correspondingspecimens, to the Royal Society on his death in 1723 at theage of 90. Leeuwenhoek had actively discouraged teachinghis methods, for reasons that are troubling today in an agewhen education is open to all. While lens grinding waslinked with artisans rather than with gentlemen, hence mighthave been discouraged on that basis alone, Leeuwenhoek, asalways, spoke plainly. In a letter to Leibnitz, he wrote ‘Totrain young people to grind lenses, and to found a sort ofschool for this purpose, I can’t see there’d be much use: becausemany students at Leyden have already been fired by my dis-coveries and my lens grinding . . . But what’s come of it?Nothing, as far as I know: because most students go there tomake money out of science, or to get a reputation in the learnedworld. But in lens grinding, and discovering things hiddenfrom our sight, these count for nought. And I’m satisfied toothat not one man in a thousand is capable of such study,because it needs much time, and spending much money; andyou must always keep on thinking about these things, if youare to get any results. And over and above all, most men arenot curious to know: nay, some even make no bones aboutsaying: What does it matter whether we know this or not?’



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[30]. Most scientists, I imagine, would see themselves as thatone man in a thousand; it is our task today to persuadeothers that it does indeed matter, not for any immediate benefit,but for the sake of curiosity and its unknowable contribution tothe sum of human knowledge and wellbeing.

The dominant use of compound microscopes over the fol-lowing centuries meant that the brief blaze of Leeuwenhoek’sdiscoveries was nearly extinguished until the great compoundmicroscope makers of the early-nineteenth century, notablyJoseph Bancks (who also produced some high-poweredsingle-lens microscopes, used by Robert Brown in his discoveryof Brownian motion and cytoplasmic streaming, and byDarwin aboard the Beagle). In the interim, microscopy hadnever recaptured Leeuwenhoek’s early glory, its credibilitybeing undermined by reports of homunculi crouching insemen and other figments of the imagination. The concept ofpreformation was called into serious question from the 1740s,beginning with Abraham Trembley’s work on the regenerationof freshwater polyps [31]. In the 1750s, Linnaeus scarcelytroubled himself with the classification of microbes; hedumped the whole lot into the phylum Vermes (‘worms’),genus Chaos (formless). The damaging accusation of seeingthings that were not there, combined with Linnaeus’s insinuatedabsence of structure, meant that few believed Leeuwenhoekcould have seen cells as small as bacteria; even the empatheticDobell struggled to conceive what magical form of lightingLeeuwenhoek must have employed to view his specimens.Only the galvanizing work of Brian J. Ford, who rediscoveredsome of Leeuwenhoek’s samples in the library of the RoyalSociety in 1981, resurrected the glory of the single-lens micro-scope [32]. Ford photographed Leeuwenhoek’s originalspecimens using one of his surviving microscopes in Utrecht,and demonstrated a remarkable resolution of less than 1 mm[33] (figure 4). That left little scope for disbelief: plainly,Leeuwenhoek really did see much of what he claimed.

So what is Leeuwenhoek’s legacy? Most of his discoverieswere forgotten, and only rediscovered in the nineteenth cen-tury, 150 years later, being then interpreted in the context ofthe newly developing cell theory, with little reference back toLeeuwenhoek himself. In this regard Leeuwenhoek’s legacy isanalogous to that of Gregor Mendel, likewise rediscovered ata time when others were exploring similar ideas. Leeuwen-hoek’s work, of course, ranged far beyond microbiology. Inall, he sent around 200 letters to the Royal Society, 112 ofwhich were published, touching on many aspects of biologyand even mineralogy. He remains the most highly publishedauthor in the journal. He is considered to be the founder ofmany fields, but none of them more important than his aston-ishing discoveries in microbiology, and none conveyed withsuch delight. Leeuwenhoek was captivated by his animalcules.‘Among all the marvels that I have discovered in nature’, hewrote, ‘these are the most marvellous of all’ [34]. His exhilara-tion in discovery, combined with a fearless and surefootedinterpretation of unknown vistas, is for me Leeuwenhoek’strue legacy. It is a spirit effervescent in many later pioneers ofmicrobiology, indeed in science more generally. And manyof the problems that beset Leeuwenhoek troubled them too.

Take the ultrastructure of cells, especially protists. Leeu-wenhoek could clearly see ‘little feet’ (cilia) and also thebudding offspring of cells, but he saw much more thanthat. I’m struck by this passage in the 1677 paper, describingan ‘egg-shaped’ animalcule (which Dobell tentatively ident-ified as the ciliate Colpidium colpoda [35]): ‘Their body did

consist, within, of 10, 12, or 14 globuls, which lay separatefrom each other. When I put these animalcula in a dry place,they then changed their body into a perfect round, andoften burst asunder, & the globuls, together with someaqueous particles, spred themselves every where about, with-out my being able to discern any other remains. Theseglobuls, which in the bursting of these creatures did flowasunder here and there, were about the bigness of the firstvery small creatures [bacteria]. And though as yet I couldnot discern any small feet in them, yet me thought, theymust needs be furnished with very many . . . ’ [1].

While the ‘globuls’ in C. colpoda were probably mostly foodvacuoles, as well as the macronucleus, Leeuwenhoek’s com-parison with bacteria leaves open the tantalizing possibilitythat he had even seen organelles such as mitochondria,which with a diameter of 0.5–1 mm would have pushed hismicroscopical resolution to the limits. Some 250 years later,this equivalence between intracellular ‘globuls’ and free-living bacteria was pursued by the early-twentieth centurypioneers of endosymbiotic theory, notably the RussianKonstantin Mereschkowski, Frenchman Paul Portier andAmerican Ivan Wallin, the latter pair independently going sofar as to argue that mitochondria could be cultivated [36].The idea of ‘symbiogenesis’ was famously ridiculed by theAmerican cell biologist E.B. Wilson, who summed up the pre-vailing attitude: ‘To many, no doubt, such speculations mayappear too fantastic for present mention in polite biologicalsociety; nevertheless, it is within the range of possibility thatthey may some day call for serious consideration’ [37]. Anotherhalf-century was to elapse before Lynn Margulis and othersdemonstrated that mitochondria and chloroplasts do indeedderive from bacterial endosymbionts [38]; and even then, notwithout a fight. I doubt that the idea of endosymbiosiswould have shocked Leeuwenhoek; nor would he have beenmuch surprised by the contemptuous disbelief of manybiologists over decades.

Another unifying theory came from biochemistry, and fit-tingly drew inspiration from Leeuwenhoek’s hometown ofDelft (described by the Earl of Leicester, once Governor-General of the Netherlands, as ‘another London almost forbeauty and fairness’). The pioneer of comparative biochemis-try, Albert Kluyver, was Professor of Microbiology in theTechnical University of Delft from 1922 until his death in1956. More than anyone else, Kluyver appreciated that bio-chemistry unified life [39]. He realized that different typesof respiration (he cites sulfate reduction, denitrification andmethanogenesis) are fundamentally equivalent, all involvingthe transfer of electrons from a donor to an acceptor. Heappreciated that all forms of respiration and fermentationare united in that they all drive growth by means of phos-phorylation. Such parallels made the startling differencesbetween cells explicable, a discovery he cherished as ‘highlyedifying to the scientific mind’ [40]. He expressed this unityin the awkward phrase ‘From elephant to butyric acid bacter-ium—it is all the same’, later paraphrased, more memorablybut without attribution, by François Jacob and JacquesMonod as ‘that old axiom ‘what is true for bacteria is alsotrue for elephants’’. Kluyver, in a seminal passage, recog-nized that the fundamental unity of biochemistry ‘opensthe way for a better appreciation of evolutionary develop-ments which have taken place in the microbial world, sincethe antithesis between the aerobic and anaerobic mode oflife has been largely removed’ [40].


eubacteria archaebacteria

eukaryotes

Figure 5. A tree of life drawn by Bill Martin in 1998, reflecting whole genomes. The tree shows the chimeric origin of eukaryotes, in which an archaeal host cellacquired bacterial endosymbionts that evolved into mitochondria; and the later acquisition of chloroplasts in Plantae. Reproduced with permission from [51].Copyright 1999 & John Wiley & Sons, Inc.


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The unity of biochemistry, then, gave the first insightsinto the evolution of the astonishing variety of ‘little animals’,which until then had remained a mystery, their provenanceas wholly unknown as in Leeuwenhoek’s time. Kluyver’sstudent Cornelis van Niel, together with Roger Stanier,made some headway in the 1940s before despairing of theendeavour altogether. By the time they published theirfamous essay ‘The concept of a bacterium’ in 1961 they nolonger cared to defend their own earlier taxonomic systems[41]; they sought only to distinguish bacteria (prokaryoticcells, lacking a nucleus) from larger eukaryotic protists,all of which have a nucleus. In this, they were remarka-bly perspicacious, commenting: ‘The differences betweeneukaryotic and prokaryotic cells are not expressed in anygross features of cellular function; they reside rather indifferences with respect to the detailed organization of the cellularmachinery’ [41]. They cite the examples of respiration andphotosynthesis, found in both eukaryotic and prokaryoticcells: ‘But in the prokaryotic cell, these metabolic unitprocesses are performed by an apparatus which alwaysshows a much smaller degree of specific organization. Infact, one can say that no unit of structure smaller than thecell in its entirety is recognizable as the site of either metabolicunit process’ [41]. This is a beautiful insight, worthy ofLeeuwenhoek himself. In eukaryotes, respiration and photo-synthesis are conducted in mitochondria and chloroplasts,respectively, and continue perfectly well in isolation fromthe rest of the cell, as all the soluble enzymes needed areconstrained within the bioenergetic membranes of theorganelle. In bacteria, by contrast, the enzymes requiredare split between the cell membrane (whether invaginatedor otherwise) and the cytosol, making the bacterium asa whole the indivisible functional unit. This distinctionapplies as much to cyanobacteria (classed as algae, not bac-teria, by Ernst Haeckel and later systematists) as to otherbacteria. Stanier and van Niel therefore argued that bacteriaare a single (monophyletic) group, all similar in their

basic plan, but insisted that any further attempts to definephylogeny were hopeless.

The timing was unfortunate. Francis Crick had alreadyadvocated the use of molecular sequences as a wonderfullysensitive phylogenetic signal, writing in 1958: ‘Biologistsshould realize that before long we shall have a subject whichmight be called ‘protein taxonomy’—the study of amino acidsequences of proteins of an organism and the comparison ofthem between species. It can be argued that these sequencesare the most delicate expression possible of the phenotypeof an organism and that vast amounts of evolutionaryinformation may be hidden away within them’ [42]. Soonafterwards, Zuckerkandl & Pauling [43] formalized the argu-ment with sequence data; and a mere two decades later,Carl Woese published his first tree of life [44]. Woese [45]was soon dismissing Stanier and van Niel as epitomising thedark ages of microbiology, when microbiologists had givenup any prospect of a true phylogeny. Woese’s tree was basedon ribosomal RNA. He showed that prokaryotes are notmonophyletic at all, but subdivide into two great domains,the bacteria and archaea. Later work, which used othermethods to ‘root’ the tree [46], portrayed the eukaryotes as a‘sister group’ to the archaea [47]. For the first time, it seemedpossible to reconstruct the evolutionary relationships betweenLeeuwenhoek’s animalcules in an evolutionary tree of life.Woese and his co-workers went so far as to argue that theterm prokaryote was obsolete, being an invalid negative defi-nition (i.e. prokaryotes are defined by the absence of a nucleus;[48]). The three domains tree is still the standard text bookview. Even so, for all its revolutionary appeal, Woese’s treeis the apotheosis of a reductionist molecular view of evolution,based on constructing trees from a single gene. It is ironic that,later in life, Woese called for a more holistic biology, whilerefusing to countenance the limitations of his single-genetree [49].

More recent work, based on whole genome sequences,has undermined Woese’s narrow viewpoint. While the



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sisterhood of archaea and eukaryotes is upheld for a core ofinformational genes—genes involved in DNA replication,transcription and translation—it is not at all true for mostother genes in eukaryotes, which are more closely related tobacteria than archaea. Woese’s iconic tree is therefore pro-foundly misleading, and should be seen strictly as a tree ofone gene only: it is not a tree of life. We cannot infer whata cell might have looked like, or how it might have lived inthe past, on the basis of its ribosomal genotype. Eukaryotesare now plainly seen to be genomic chimeras, apparentlyformed in a singular endosymbiosis between an archaealhost cell and a bacterium around 1.5 billion years ago [50].This chimerism cannot be depicted on a normal branchingphylogenetic tree, because endosymbiosis involves fusionof branches, not bifurcation, producing instead a strikingcomposite tree, depicted beautifully (and presciently, as thisis still accurate) by Bill Martin in 1998 [51] (figure 5). BillMartin and I have since argued that the singular endosym-biosis at the origin of eukaryotes, which gave rise tomitochondria, increased the energy available per gene ineukaryotic cells by a breath-taking three to five orders of mag-nitude [52]. That overcame the pervasive energetic constraintsfaced by bacteria, enabling a massive expansion in cellvolume and genome size, and permitting the evolution ofmany eukaryotic traits from the nucleus to sex and phagocyto-sis (all of which were first reported by Leeuwenhoek himself).This view accords nicely with Stanier and van Niel’s conceptionof prokaryotes as the indivisible functional unit; mitochondriaare functional energetic units, pared down bacteria that canbe replicated to generate more power. It might be that eukar-yotes had to evolve by way of an endosymbiosis, for thesebioenergetic reasons.

Even in the absence of endosymbiosis, the idea of a truephylogenetic tree of life is undermined by the prevalence of lat-eral gene transfer in both bacteria and archaea. Informationalgenes, including ribosomal RNA, are generally inherited verti-cally, giving a robust phylogenetic signal, but such genesaccount for barely 1% of a bacterial genome, and much of therest is passed around between cells by lateral gene transfer,confounding deep phylogenetic signals. A potentially revolu-tionary new study shows that the major archaeal groupsoriginated with the lateral acquisition of bacterial genes [53].Ironically, the unity of biochemistry—Kluyver’s edifyingguide to evolution—is the root problem: the universality ofthe genetic code, intermediary metabolism and energy conser-vation (e.g. the shared mechanism of respiration) means thatgenes are an exchangeable currency, and facilitate adaptation

to the endless variety of external conditions. Again, the linkbetween the ribosomal genotype of a prokaryotic cell and itsphenotype—the way it makes a living—is forever changing.Ford Doolittle notes that pervasive genetic chimerism meansthat ‘no hierarchical universal classification can be taken asnatural’ [54]; the universal tree of life is a human foible andnot a true representation of the real world. As Doolittleobserves, ‘Biologists might rejoice in and explore, rather thanregret or attempt to dismiss, the creative evolutionary role oflateral gene transfer’ [54]. The tree of life promises a hierarchi-cal order, and takes authority from Darwin himself, but inmicrobes at least it is not sustained by the very geneticsequences that made such phylogeny possible. ‘Early evolutionwithout a tree of life’ [55] might seem an alarming vista tomany, but Leeuwenhoek would surely have felt at home. Hewas happiest without a compass.

Perhaps that, more than anything else, is the lesson westill need to learn from Leeuwenhoek today. There is adanger of complacency in biology, a feeling that the immensecomputational power of the modern age will ultimatelyresolve the questions of biology, and medical research morebroadly. But pathophysiology stems from physiology, andphysiology is a product of evolution, largely at the level ofcells. The eukaryotic cell seems to have arisen in a singularendosymbiosis between prokaryotes, and eukaryotes sharea large number of basic traits, few of which are known in any-thing like the same form in bacteria or archaea. We know ofno surviving evolutionary intermediates between prokar-yotes and eukaryotes. We know almost nothing aboutwhich factors drove the evolution of many basal eukaryotictraits, from the nucleus to meiosis and sex, to cell death—traits first observed by Leeuwenhoek. Why did meiosis andsex arise from lateral gene transfer in bacteria? Why did thenucleus evolve in eukaryotes but not in bacteria or archaea?What prevents bacteria from engulfing other cells by phago-cytosis? There is no agreement on the answers to thesequestions, nor more broadly to a question that might easilyhave been asked by Leeuwenhoek himself—why is lifethe way it is? Some of us have argued that eukaryotic evol-ution is explicable in terms of the detailed mechanisms ofenergy conservation, with an allied requirement for endo-symbiosis leading to conflict and coadaptation betweenendosymbionts and their host cells [56]. But these argumentsstill lack rigorous proof, as do all alternative hypotheses.In the meantime, we have at best an unreliable map ofthe land that enchanted Leeuwenhoek. We should rejoiceand explore.

Author profile

Nick Lane is a Reader in Evolutionary Biochemistry in the Department of Genetics, Evolutionand Environment at University College London. His research is on the role of bioenergetics inthe origin of life and the early evolution of cells, focusing on the importance of endosymbiosisand cellular structure in determining the course of evolution. Nick was awarded the inauguralUCL Provost’s Venture Research Fellowship in 2009, the BMC Research Award for Genetics,Genomics, Bioinformatics and Evolution in 2011, and the Biochemical Society Award for 2015.He leads the UCL Research Frontiers Origins of Life programme and was a founding memberof the UCL Consortium for Mitochondrial Research. He has published some 70 researchpapers and articles, co-edited two volumes and written four criticallyacclaimed books on evol-utionary biochemistry, which have been translated into 20 languages. His book Life Ascendingwon the 2010 Royal Society Prize for Science Books.


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References


370:20140344

1. Leewenhoeck A. 1677 Observation, communicatedto the publisher by Mr. Antony van Leewenhoeck,in a Dutch letter of the 9 Octob. 1676 hereEnglish’d: concerning little animals by him observedin rain-well-sea and snow water; as also inwater wherein pepper had lain infused. Phil. Trans.12, 821 – 831. (doi:10.1098/rstl.1677.0003)

2. Letter from Constantijn Huygens to Robert Hooke, 8August 1673, translated in Dobell C. 1958 Antonyvan Leeuwenhoek and his little animals, p. 43.New York, NY: Russell and Russell.

3. Hooke R. 1665 Micrographia. London, UK: Jo. Martynand Ja. Allestry, printers to the Royal Society.

4. Letter from Leeuwenhoek to Oldenburg, 7September 1674, translated in Dobell C. 1958Antony van Leeuwenhoek and his little animals,pp. 109 – 110. New York, NY: Russell and Russell.

5. Letter from Leeuwenhoek to Hooke, 12 November1680, translated in Dobell C. 1958 Antony vanLeeuwenhoek and his little animals, p. 200.New York, NY: Russell and Russell.

6. Dobell C. 1958 Antony van Leeuwenhoek and his littleanimals, p. 113. New York, NY: Russell and Russell.

7. Ruestow EG. 2004 The microscope in the Dutch Republic.Cambridge, UK: Cambridge University Press.

8. Letter from Leeuwenhoek to Constantijn Huygens, 7November 1676, translated in Dobell C. 1958 Antonyvan Leeuwenhoek and his little animals, p. 116.New York, NY: Russell and Russell.

9. Letter from Leeuwenhoek to Oldenburg, 15 August1673, translated in Dobell C. 1958 Antony vanLeeuwenhoek and his little animals, p. 42.New York, NY: Russell and Russell.


11. Letter from Leeuwenhoek to Oldenburg, 23 March1677, translated in Dobell C. 1958 Antony vanLeeuwenhoek and his little animals, p. 171.New York, NY: Russell and Russell.

12. Letter from Leeuwenhoek to Constantijn Huygensand Hooke, 20 May 1679, translated in Dobell C.1958 Antony van Leeuwenhoek and his littleanimals, p. 188. New York, NY: Russell and Russell.

13. Dobell C. 1958 Antony van Leeuwenhoek and hislittle animals, p. 141, note 1. New York, NY: Russelland Russell.

14. Letter from Leeuwenhoek to Oldenburg, 5 October1677, translated in Dobell C. 1958 Antony vanLeeuwenhoek and his little animals, p. 174.New York, NY: Russell and Russell.


16. van Lewuenhoeck A. 1702 Part of a Letter from MrAntony van Leeuwenhoek, F. R. S. concerning greenweeds growing in water, and some animalculafound about them. Phil. Trans. 23, 1304 – 1311.(doi:10.1098/rstl.1702.0042)

17. Leewenhoeck A. 1684 An Abstract of a Letter fromMr. Anthony Leewenhoeck at Delft, dated Sep. 17.1683. containing some microscopical observations,

about animals in the scurf of the teeth, thesubstance call’d worms in the nose, the cuticulaconsisting of scales. Phil. Trans. 14, 568 – 574.(doi:10.1098/rstl.1684.0030)

18. Shapin S. 1994 A social history of truth. Chicago, IL:University of Chicago Press.

19. Letter from Christiaan Huygens to Oldenburg, 30January 1675, translated in Dobell C. 1958 Antonyvan Leeuwenhoek and his little animals, p. 172.New York, NY: Russell and Russell.

20. Hooke R. 1678 Lectures and collections;Microscopium. London, UK: J Martyn, Printer to theRoyal Society.

21. Letter from Hooke to Leeuwenhoek, 1 December1677, translated in Dobell C. 1958 Antony vanLeeuwenhoek and his little animals, p. 183.New York, NY: Russell and Russell.

22. Ford BJ. 1985 Single lens. New York, NY: Harper andRow.

23. Gest H. 2004 The discovery of microorganisms byRobert Hooke and Antoni van Leeuwenhoek,Fellows of the Royal Society. Notes Rec. R. Soc. Lond.58, 187 – 201. (doi:10.1098/rsnr.2004.0055)

24. Dobell C. 1958 Antony van Leeuwenhoek and hislittle animals, pp. 163 – 164. New York, NY: Russelland Russell.

25. Oeuvres Compl de Chr. Huygens Vol VIII (1899) andXIII (1916).

26. Preface to Hooke R. 1665 Micrographia. London, UK:Royal Society.

27. Ford BJ. 1985 Single lens: the story of the simplemicroscope, p. 40. London, UK: William Heinemann.

28. Ford BJ. 2005 The discovery of Giardia. TheMicroscope 53, 147 – 153.

29. Ford BJ. 2007 Antony van Leeuwenhoek’smicroscope and the discovery of Giardia. Microscopyand Analysis 21, 5 – 7.

30. Letter from Leeuwenhoek to Leibnitz, 28 September1725, translated in Dobell C. 1958 Antony vanLeeuwenhoek and his little animals, p. 325.New York, NY: Russell and Russell.

31. Lenhoff SG, Lenhoff HM. 1986 Hydra and the birthof experimental biology, 1744: Abraham Trembley‘smemoires concerning the polyps. Pacific Grove, CA:Boxwood Press.

32. Ford BJ. 1981 The van Leeuwenhoek specimens.Notes Rec. R. Soc. Lond. 36, 37 – 59.

33. Ford BJ. 1991 The Leeuwenhoek legacy. London, UK:Biopress, Briston and Farrand Press.

34. Letter from Leeuwenhoek to Oldenburg, 9 October1676, translated in Dobell C. 1958 Antony vanLeeuwenhoek and his little animals, p. 144.New York, NY: Russell and Russell.

35. Dobell C. 1958 Antony van Leeuwenhoek and hislittle animals, pp. 134 – 135 notes. New York, NY:Russell and Russell.

36. Sapp J. 1994 Evolution by association: a history ofsymbiosis. New York, NY: Oxford University Press.

37. Wilson EB. 1925 The Cell in development andheredity. New York, NY: Macmillan.

38. Sagan L. 1967 On the origin of mitosing cells.J. Theor. Biol. 14, 225 – 274. (doi:10.1016/0022-5193(67)90079-3)

39. Woods DD. 1957 Albert Jan Kluyver. 1888 – 1956.Biogr. Mems. Fell. R. Soc. 3, 109 – 122. (doi:10.1098/rsbm.1957.0008)

40. Kluyver AJ. 1947 Three decades of progress inmicrobiology. Antonie van Leeuwenhoek 13, 1 – 20.(doi:10.1007/BF02272745)

41. Stanier RY, van Niel CB. 1961 The concept of abacterium. Arch. Microbiol. 42, 17 – 35. (doi:10.1007/BF00425185)

42. Crick FHC. 1958 The biological replication ofmacromolecules. Symp. Soc. Exp. Biol. 12, 138 – 163.

43. Zuckerkandl E, Pauling L. 1962 Molecular disease,evolution and genetic heterogeneity. In Horizons inbiochemistry (eds M Kasha, B Pullman), pp. 189 –225. New York, NY: Academic Press.

44. Woese CR, Fox GE. 1977 Phylogenetic structure ofthe prokaryotic domain: the primary kingdoms.Proc. Natl Acad. Sci. USA 74, 5088 – 5090. (doi:10.1073/pnas.74.11.5088)

45. Woese CR. 1994 There must be a prokaryote somewhere:microbiology’s search for itself. Microbiol. Rev. 58, 1 – 9.

46. Iwabe N, Kuma K, Hasegawa M, Osawa S, Miyata T. 1989Evolutionary relationship of archaebacteria, eubacteria,and eukaryotes inferred from phylogenetic trees ofduplicated genes. Proc. Natl Acad. Sci. USA 86,9355 – 9359. (doi:10.1073/pnas.86.23.9355)

47. Woese C, Kandler O, Wheelis ML. 1990 Towards anatural system of organisms: proposal for the domainsArchaea, Bacteria, and Eucarya. Proc. Natl Acad. Sci. USA87, 4576 – 4579. (doi:10.1073/pnas.87.12.4576)

48. Sapp J. 2005 The prokaryote-eukaryote dichotomy:meanings and mythology. Microbiol. Mol. Biol. Rev. 69,292 – 305. (doi:10.1128/MMBR.69.2.292-305.2005)

49. Woese CR. 2004 A new biology for a new century.Microbiol. Mol. Biol. Rev. 68, 173 – 186. (doi:10.1128/MMBR.68.2.173-186.2004)

50. Williams TA, Foster PG, Cox CJ, Embley TM. 2013 Anarchaeal origin of eukaryotes supports only twoprimary domains of life. Nature 504, 231 – 236.(doi:10.1038/nature12779)

51. Martin W. 1999 Mosaic bacterial chromosomes: achallenge en route to a tree of genomes. Bioessays21, 99 – 104. (doi:10.1002/(SICI)1521-1878(199902)21:2,99::AID-BIES3.3.0.CO;2-B)

52. Lane N, Martin W. 2010 The energetics of genomecomplexity. Nature 467, 929 – 934. (doi:10.1038/nature09486)

53. Nelson-Sathi S et al. 2015 Origins of major archaealclades correspond to gene acquisitions from bacteria.Nature 517, 77 – 80. (doi:10.1038/nature13805)

54. Doolittle WF. 1999 Phylogenetic classification andthe universal tree. Science 284, 2124 – 2128.(doi:10.1126/science.284.5423.2124)

55. Martin WF. 2011 Early evolution without a tree of life.Biol. Direct. 6, 36. (doi:10.1186/1745-6150-6-36)

56. Lane N. 2015 The vital question: energy, evolution andthe origins of complex life. New York, NY: WW Norton.

http://dx.doi.org/10.1098/rstl.1677.0003http://dx.doi.org/10.1098/rstl.1702.0042http://dx.doi.org/10.1098/rstl.1684.0030http://dx.doi.org/10.1098/rsnr.2004.0055http://dx.doi.org/10.1016/0022-5193(67)90079-3http://dx.doi.org/10.1016/0022-5193(67)90079-3http://dx.doi.org/10.1098/rsbm.1957.0008http://dx.doi.org/10.1098/rsbm.1957.0008http://dx.doi.org/10.1007/BF02272745http://dx.doi.org/10.1007/BF00425185http://dx.doi.org/10.1007/BF00425185http://dx.doi.org/10.1073/pnas.74.11.5088http://dx.doi.org/10.1073/pnas.74.11.5088http://dx.doi.org/10.1073/pnas.86.23.9355http://dx.doi.org/10.1073/pnas.87.12.4576http://dx.doi.org/10.1128/MMBR.69.2.292-305.2005http://dx.doi.org/10.1128/MMBR.68.2.173-186.2004http://dx.doi.org/10.1128/MMBR.68.2.173-186.2004http://dx.doi.org/10.1038/nature12779http://dx.doi.org/10.1002/(SICI)1521-1878(199902)21:2%3C99::AID-BIES3%3E3.0.CO;2-Bhttp://dx.doi.org/10.1002/(SICI)1521-1878(199902)21:2%3C99::AID-BIES3%3E3.0.CO;2-Bhttp://dx.doi.org/10.1002/(SICI)1521-1878(199902)21:2%3C99::AID-BIES3%3E3.0.CO;2-Bhttp://dx.doi.org/10.1002/(SICI)1521-1878(199902)21:2%3C99::AID-BIES3%3E3.0.CO;2-Bhttp://dx.doi.org/10.1002/(SICI)1521-1878(199902)21:2%3C99::AID-BIES3%3E3.0.CO;2-Bhttp://dx.doi.org/10.1002/(SICI)1521-1878(199902)21:2%3C99::AID-BIES3%3E3.0.CO;2-Bhttp://dx.doi.org/10.1002/(SICI)1521-1878(199902)21:2%3C99::AID-BIES3%3E3.0.CO;2-Bhttp://dx.doi.org/10.1038/nature09486http://dx.doi.org/10.1038/nature09486http://dx.doi.org/10.1038/nature13805http://dx.doi.org/10.1126/science.284.5423.2124http://dx.doi.org/10.1186/1745-6150-6-36http://rstb.royalsocietypublishing.org/

The unseen world: reflections on Leeuwenhoek (1677) 'Concerning little animals'Author profileReferences

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